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Swine-origin influenza A virus head protein and preparation method and application thereof

A technology of influenza virus and protein, which is applied in the head protein of porcine type A influenza virus and its preparation and application, can solve the problems of high production cost and long production cycle, reduce production cost, shorten production time, and ensure biological safety effect

Active Publication Date: 2014-01-08
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It faces the problem of long production cycle and high production cost in the production process, and needs a large amount of SPF chicken embryos in the production process

Method used

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  • Swine-origin influenza A virus head protein and preparation method and application thereof
  • Swine-origin influenza A virus head protein and preparation method and application thereof
  • Swine-origin influenza A virus head protein and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Influenza virus protein

[0029] The amino acid composition of the influenza protein is shown in SEQ ID NO.1.

[0030] SEQ ID NO.1:APLHLGKCNIAGWILGNPECESLSTASSWSYIVETPSSDNGTCYPGDFIDYEELREQLSSVSSFERFEIFPKTSSWPNHDSNKGVTAACPHAGAKSFYKNLIWLVKKGNSYPKLSKSYINDKGKEVLVLWGIHHPSTSADQQSLYQNADTYVFVGSSRYSKKFKPEIAIRPKVRDQEGRMNYYWTLVEPGDKITFEATGNLVVPRYAFAMERN

[0031] The amino acid composition of the influenza virus protein can also be: a protein whose amino acid sequence in the protein shown in SEQ ID NO.1 is substituted, deleted or added with one or several amino acids and has influenza protein antigenicity, such as SEQ ID NO.2 :

[0032] APLQLGKCNIAGWLLGNPECDLLLTASSWSYIVETSNSENGTCYPGDFIDYEELREQLSSVSSFEKFEIFPKTSSWPNHETTKGVTAACSYAGASSFYRNLLWLTKKGSSYPKLSKSYVNNKGKEVLVLWGVHHPPTGTDQQSLYQNADAYVSVGSSKYNRRFTPEIAARPKVRDQAGRMNYYWTLLEPGDTITFEATGNLIAPWYAFALN;或SEQ ID NO.3:

[0033] APLQLGKCNIAGWLLGNPECDPLLPVRSWSYIVETPNSENGICYPGDFIDYEELREQLSSVSSFERFEIFPKESSWPNHNTTKGVTAACSHAGKSSFYRNLL...

Embodiment 2

[0034] Example 2 Preparation of influenza virus protein

[0035] 1. Obtaining the target gene

[0036] The influenza virus A / California / 04 / 2009(H1N1), GenBank: FJ966082.1 selected in this experiment, was artificially synthesized according to the sequence of GenBank: FJ966082.1 (this synthesis was provided by Shanghai Sangon Bioengineering Technology Service Co., Ltd.) Acid sequence SEQ ID NO.4,

[0037] gccccatt gcatttgggt aaatgtaaca ttgctggctg gatcctggga aatccagagt gtgaatcact ctccacagca agctcatggt cctacattgt ggaaacacct agttcagaca atggaacgtg ttacccagga gatttcatcg attatgagga gctaagagag caattgagct cagtgtcatc atttgaaagg tttgagatat tccccaagac aagttcatgg cccaatcatg actcgaacaa aggtgtaacg gcagcatgtc ctcatgctgg agcaaaaagc ttctacaaaa atttaatatg gctagttaaa aaaggaaatt catacccaaa gctcagcaaa tcctacatta atgataaagg gaaagaagtc ctcgtgctat ggggcattca ccatccatct actagtgctg accaacaaag tctctatcag aatgcagata catatgtttt tgtggggtca tcaagataca gcaagaagtt caagccggaa atagcaataa gacccaaagt gagggat...

Embodiment 3

[0062] Example 3 Protein Animal Test

[0063] H1HA head region protein immunogenicity experiment:

[0064] Experimental animals: 24 SPF chickens, each group of 6 divided into 4 groups, namely: blank control group, adjuvant control group, 50 μg antigen amount group, 100 μg antigen amount group

[0065] Reagents: complete Freund's adjuvant and incomplete Freund's adjuvant were purchased from Sigma

[0066] Methods: 50 μg and 100 μg of purified H1HA head region recombinant protein were mixed with Freund's complete adjuvant for intramuscular injection immunization, and 3 weeks later, the corresponding antigen was mixed with incomplete Freund's adjuvant for booster immunization. At the same time, a blank control group and an adjuvant control group were set up. Chicken serum was collected before immunization and 10 days after the last immunization. On the 21st day after the last immunization, 50 μL containing 10 6 EID 50 The dilution of A / California / 04 / 2009 (H1N1...

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Abstract

The invention discloses a swine-origin influenza A virus head protein and a preparation method and application thereof. The protein has an amino acid sequence shown as SEQ ID NO.1. By the method, the yield of hemagglutinin (HA) in influenza virus can be improved, and more than 200 milligrams of purified HA protein can be obtained in each two liters of culture medium; a batch of purified proteins can be obtained only after 5 to 6 days, and production time is shortened; in addition, the influenza virus is not used, so the biological safety of a vaccine can be ensured, and the danger of spreading poison is avoided; a chick embryo is not required to be used for amplifying the virus, so production cost is greatly reduced; and the protein has a good application prospect.

Description

technical field [0001] The invention relates to an influenza virus protein and its preparation method and application, in particular to a highly expressed functional porcine type A influenza head protein. Background technique [0002] Influenza (abbreviated as influenza) (influenza) is a kind of acute infectious disease caused by influenza virus caused by humans, poultry and animals. Influenza viruses belong to the family Orthomyxoviridae. According to the characteristics of viral nucleocapside protein (NP) and matrix protein (matrix protein), influenza viruses are divided into three types: A (A), B (B) and C (C). Influenza A viruses can be further divided into different subtypes according to the antigenicity of the hemagglutinin (HAmagglutinin, HA) and neuraminidase (neuraminidase, NA) proteins on the surface of virus particles. There are 16 HA subtypes and 9 NA subtypes of influenza A virus found so far. Influenza A virus was discovered in 1901. However, human influenz...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/11C12R1/19
Inventor 高福宣春玲
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI